The development of a general method for studying the sequence specificity of small ligands which bind to DNA is proposed. The method involves partial digest of a drug ligated, singly end-labeled (32P), DNA restriction fragment using a nonspecific endonuclease. The DNA fragments produced in the digest along with the base specific reaction products of the Maxam-Gilbert sequencing method, are separated using electrophoresis in a polyacrylamide sequencing gel. Autoradiography and microdensitometry allows quantitative comparison of the digest patterns obtained in the presence of the drug with those obtained in its absence. The regions of protection from nuclease attack, allow direct visualization of the binding sequence of the drug as well as the determination of thermodynamic properties associated with the drug DNA contact. In addition to examining the relationship between a drug's structure and its sequence specificity for linear DNA's, the method will be adapted to the study of drug-DNA interactions using supercoiled DNA and chromatin as binding substrates.